HiDi 2x PCR Master Mix

HiDi 2x PCR Master Mix

  • HiDi 2x PCR Master Mix - ready to use mix simplifies your PCR setup for single nucleotide discrimination. Only target-specific primers and template need to be added as the mix contains all components for a successful and reliable PCR. This ensures reproducible results, significantly reduces set-up times and the risk of pipetting errors.  

    HiDi is the best choice for highly selective PCRs, such as allele-specific PCR, HLA genotyping, analysis of single CpG methylation sites or the detection of mutations in a high background of wild-type sequences as shown by benchmarking data with other well-known enzymes on the market. HiDi DNA polymerase efficiently discriminates mismatched primer-template complexes and only produces specific amplicons in case of perfectly matched primer pairs. 

  • This mix contains an engineered DNA polymerase in an optimzed reaction buffer and ultrapure dNTPs. HiDi DNA polymerase is hotstart-formulated with an aptamer. Temperatures above 50-55°C cause the aptamer’s secondary structure to melt and will set-free the polymerase. 

    HiDi DNA polymerase can also be used for real-time cycling, when adding a suitable real-time dye, for example GreenDye (#2000).

    However, this mix can not be used for hydrolysis-based probes (e.g. TaqMan) as the 5'-3' nuclease is not active. For this application see HiDi Taq polymerase (#9201).

  • This product is shipped on cool packs. It is recommended to store the product upon arrival at -20°C.

  • HiDi 2x PCR Master Mix is tested for successful ASA performance detecting a genomic SNP (rs72921001) in HeLa genomic DNA. PCR products were subsequently analysed on a 2.5% agarose gel. Specific product was visualized by ethidium bromide staining at the correct amplicon length of 109 bp for the matched primer. In case of mismatch primer no product formation was observed after 45 cycles. The activity of HiDi DNA polymerase was monitored and adjusted to a specific DNA polymerase activity using an artificial DNA template and a DNA primer. Enzyme concentration was determined by protein-specific staining. Please inquire more information at info@mypols.de for the lot-specific concentration. No contamination was detected in standard test reactions.

Example Primer Design

Matching vs. mismatching nucleotide is placed at the 3'-end of the primer for best discrimination results.

Example Results - There´s no accounting for taste

Cilantro: some people love it in their food, some hate it. Here we are detecting a genomic SNP (rs72921001) in HeLa genomic DNA. This SNP is reported to be close to a number of genes coding for olfactory receptors. (Reference: Eriksson N. et al. (2012), "A genetic variant near olfactory receptor genes influences cilantro preference.")

Considering, that only the C-allele specific primer is extended and yielding in a specific amplicon, we can conclude a genetic predisposition in disliking cilantro, as this SNP is significantly associated with detecting a soapy taste to cilantro.  

Allele-specific PCRs were performed from 1 ng/µl of HeLa gDNA in the presence of a realtime dye, indicating the amplification of the C-allele specific primer only. The A-allele specific primer is discriminated, thus not amplified up to 50 cycles.

PCR products were subsequently analysed on a 2.5% agarose gel. Specific product is visualized by ethidium bromide staining at the amplicon length of 109 bp.

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